Time course changes to structural, mechanical and material properties of bone in rats after complete spinal cord injury

Abstract

Objective: Characterise the spatiotemporal trabecular and cortical bone responses to complete spinal cord injury (SCI) in young rats.

Methods: 8-week-old male Wistar rats received T9-transection SCI and were euthanised 2-, 6-, 10- or 16-weeks post-surgery. Outcome measures were assessed using micro-computed tomography, mechanical testing, serum markers and Fourier-transform infrared spectroscopy.

Results: The trabecular and cortical bone responses to SCI are site-specific. Metaphyseal trabecular BV/TV was 59% lower, characterised by fewer and thinner trabeculae at 2-weeks post-SCI, while epiphyseal BV/TV was 23% lower with maintained connectivity. At later-time points, metaphyseal BV/TV remained unchanged, while epiphyseal BV/TV increased. The total area of metaphyseal and mid-diaphyseal cortical bone were lower from 2-weeks and between 6- and 10-weeks post-SCI, respectively. This suggested that SCI-induced bone changes observed in the rat model were not solely attributable to bone loss, but also to suppressed bone growth. No tissue mineral density differences were observed at any time-point, suggesting that decreased whole-bone mechanical properties were primarily the result of changes to the spatial distribution of bone.

Conclusion: Young SCI rat trabecular bone changes resemble those observed clinically in adult and paediatric SCI, while cortical bone changes resemble paediatric SCI only.

Keywords: Bone Mechanical Properties; Fourier-transform Infrared Spectroscopy; Osteoporosis; Spinal Cord Transection; microCT.

Figure 1

Representative µCT-based images of metaphyseal trabecular architecture and mean morphometric outcome measures for 2-, 6-, 10- and 16-weeks post-surgery SCI and SHAM groups (all n=8). Data shown as mean ± SD. *, ** and *** indicate p<0.05, p<0.01 and p<0.001, respectively, for SCI versus SHAM at the same post-surgical timepoint.

Figure 2

Representative µCT-based images of epiphyseal trabecular architecture and mean morphometric outcome measures for 2-, 6-, 10- and 16-weeks post-surgery SCI and SHAM groups (all n=8). Data shown as mean ± SD. *, ** and *** indicate p<0.05, p<0.01 and p<0.001, respectively, for SCI versus SHAM at the same post-surgical timepoint.

Figure 3

Representative µCT-based images of distal metaphyseal cortical bone (81-85% bone length) and mean morphometric outcome measures and tissue mineral density for 2-, 6-, 10- and 16-weeks post-surgery SCI and SHAM groups (all n=8). Data shown as mean ± SD. *, ** and *** indicate p<0.05, p<0.01 and p<0.001, respectively, for SCI versus SHAM at the same post-surgical timepoint.

Figure 4

Representative µCT-based images of mid-diaphyseal cortical bone (58-62% bone length) and mean morphometric outcome measures and tissue mineral density for 2-, 6-, 10- and 16-weeks post-surgery SCI and SHAM groups (all n=8). Data shown as mean ± SD. *, ** and *** indicate p<0.05, p<0.01 and p<0.001, respectively, for SCI versus SHAM at the same post-surgical timepoint.

Figure 5

Three-point bend-determined whole-bone and material-level mechanical properties for 2-, 6-, 10- and 16-weeks post-surgery SCI and SHAM groups (all n=8). Whole-bone mechanical properties were adjusted for body mass using the linear regression method. Data shown as mean ± SD. *, ** and *** indicate p<0.05, p<0.01 and p<0.001, respectively, for SCI versus SHAM at the same post-surgical timepoint.

Figure 6

Torsion test-determined whole-bone and material-level mechanical properties for 2-, 6-, 10- and 16-weeks post-surgery SCI and SHAM groups (all n=8). Whole-bone mechanical properties were adjusted for body mass using the linear regression method. Data shown as mean ± SD. *, ** and *** indicate p<0.05, p<0.01 and p<0.001, respectively, for SCI versus SHAM at the same post-surgical timepoint.

Figure 7

Serum bone turnover marker levels for 2-, 6-, 10- and 16-weeks post-surgery SCI and SHAM groups (all n=8). Data shown as mean ± SD.* and ** indicate p<0.05 and p<0.01, respectively for SCI versus SHAM at the same post-surgical timepoint. A) bone formation marker P1NP. B) bone resorption marker CTX-I.

Online Resource 1

Global 2D morphometric analysis of A) trabecular bone area fraction for 60 to 100% bone length and B) cortical area for 40 to 85% bone length for 2- and 16-week post-surgery SCI and SHAM rats. Trabecular and cortical VOIs used for subsequent 3D are superimposed on distributions. Data shown as mean ± SE for every 1% bone length.

Online Resource 2

Schematics to represent location, measurement approach and collected FTIR spectra.

Online Resource 3

Body mass with time post-surgery for SCI and SHAM rats. Data shown as mean ± SE. * and ** indicate p < 0.05 and p < 0.01, respectively, for SCI versus SHAM at the same post-surgical timepoint.

Online Resource 4

Sublesional bone wet masses and muscle mass for SCI and SHAM rats at 2-, 6-, 10- and 16-weeks post-surgery for the right hindlimb. A) Femur wet mass, B) Tibia wet mass and C) gastrocnemius (both medial and lateral heads) muscle mass. Data shown as mean ± SE. *** and **** indicate p < 0.001 and p<0.0001, respectively, for SCI versus SHAM at the same post-surgical timepoint.

Online Resource 5

Supralesional bone wet mass and muscle masses for SCI and SHAM rats at 2-, 6-, 10- and 16-weeks post-surgery for the right forelimb. A) Tricep mass, B) Bicep mass and C) Humeri wet mass. Data shown as mean ± SE. * and ** indicate p<0.05 and p<0.01, respectively, for SCI versus SHAM at the same post-surgical timepoint.

Online Resource 6

Femoral length for SCI and SHAM rats at 2-, 6-, 10- and 16-weeks post-surgery. Data shown as mean ± SE.